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Innate immune responses of equine monocytes cultured in equine platelet lysate

Naskou, MC;Norton, NA;Copland, IB;Galipeau, J;Peroni, JF;

Platelet lysate (PL) has been extensively used for the laboratory expansion of human mesenchymal stem cells (MSC) in order to avoid fetal bovine serum (FBS) which has been associated with immune-mediated host reactions and transmission of bovine-origin microbial contaminants. Before suggesting the routine use of PL for MSC culture, we wanted to further investigate whether PL alone might trigger inflammatory responses when exposed to reactive white blood cells such as monocytes. Our objectives were to evaluate the inflammatory profile of equine monocytes cultured with equine PL (ePL) and to determine if ePL can modulate the expression of inflammatory cytokines in lipopolysaccharide (LPS)-stimulated monocytes. In a first experiment, equine monocytes were isolated and incubated with donor horse serum (DHS), FBS, six individual donors ePL or pooled ePL from all horses. In a second experiment, monocytes were stimulated with E. coli LPS in the presence of 1, 5 or 10% DHS and/or pooled ePL. After 6 hours of incubation, cell culture supernatants were assayed via ELISA for production of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF) and Interleukin 1 (IL-1) as well as for the anti-inflammatory Interleukin 10 (IL-10). Equine monocytes incubated with pooled ePL produced significantly less TNF- and significantly more IL-10 than monocytes incubated in FBS. Even though not significant, ePL appeared to also downregulate monocytes production of IL-1. The second experiment showed that pooled ePL added to LPS-stimulated equine monocytes resulted in a significant reduction in TNF- and IL-1 production. IL-10 production was not significantly upregulated by the addition of ePL to LPS-stimulated monocytes. Finally, the addition of ePL to LPS-stimulated monocytes in the presence of various concentrations of DHS resulted to statistically significant decrease of TNF- and IL-1 compared to the control groups. This is the first study to demonstrate that ePL suppresses the release of pro-inflammatory cytokines from stimulated equine monocytes. These results encourage further exploration of PL as a homologous media substitute for FBS but also opens the possibility of investigating its use as means to suppress cell-mediated inflammation.